Size matters: Aerobic methane oxidation in sediments of shallow thermokarst lakes.

Shallow thermokarst lakes are important sources of greenhouse gases (GHGs) such as methane (CH4 ) and carbon dioxide (CO2 ) resulting from continuous permafrost thawing due to global warming. Concentrations of GHGs dissolved in water typically increase with decreasing lake size due to coastal abrasion and organic matter delivery. We hypothesized that (i) CH4 oxidation depends on the natural oxygenation gradient in the lake water and sediments and increases with lake size because of stronger wind-induced water mixing; (ii) CO2 production increases with decreasing lake size, following the dissolved organic matter gradient; and (iii) both processes are more intensive in the upper than deeper sediments due to the in situ gradients of oxygen (O2 ) and bioavailable carbon. We estimated aerobic CH4 oxidation potentials and CO2 production based on the injection of 13 C-labeled CH4 in the 0-10 cm and 10-20 cm sediment depths of small (~300 m2 ), medium (~3000 m2 ), and large (~106 m2 ) shallow thermokarst lakes in the West Siberian Lowland. The CO2 production was 1.4-3.5 times stronger in the upper sediments than in the 10-20 cm depth and increased from large (158 ± 18 nmol CO2 g-1 sediment d.w. h-1 ) to medium and small (192 ± 17 nmol CO2 g-1 h-1 ) lakes. Methane oxidation in the upper sediments was similar in all lakes, while at depth, large lakes had 14- and 74-fold faster oxidation rates (5.1 ± 0.5 nmol CH4 -derived CO2 g-1 h-1 ) than small and medium lakes, respectively. This was attributed to the higher O2 concentration in large lakes due to the more intense wind-induced water turbulence and mixing than in smaller lakes. From a global perspective, the CH4 oxidation potential confirms the key role of thermokarst lakes as an important hotspot for GHG emissions, which increase with the decreasing lake size.

Temperature sensitivity of anaerobic methane oxidation versus methanogenesis in paddy soil: Implications for the CH4 balance under global warming.

The global methane (CH4 ) budget is based on a sensitive balance between methanogenesis and CH4 oxidation (aerobic and anaerobic). The response of these processes to climate warming, however, is not quantified. This largely reflects our lack of knowledge about the temperature sensitivity (Q10 ) of the anaerobic oxidation of CH4 (AOM)-a ubiquitous process in soils. Based on a 13 CH4 labeling experiment, we determined the rate, Q10 and activation energy of AOM and of methanogenesis in a paddy soil at three temperatures (5, 20, 35°C). The rates of AOM and of methanogenesis increased exponentially with temperature, whereby the AOM rate was significantly lower than methanogenesis. Both the activation energy and Q10 of AOM dropped significantly from 5-20 to 20-35°C, indicating that AOM is a highly temperature-dependent microbial process. Nonetheless, the Q10 of AOM and of methanogenesis were similar at 5-35°C, implying a comparable temperature dependence of AOM and methanogenesis in paddy soil. The continuous increase of AOM Q10 over the 28-day experiment reflects the successive utilization of electron acceptors according to their thermodynamic efficiency. The basic constant for Q10 of AOM was calculated to be 0.1 units for each 3.2 kJ mol-1 increase of activation energy. We estimate the AOM in paddy soils to consume 2.2~5.5 Tg CH4 per year on a global scale. Considering these results in conjunction with literature data, the terrestrial AOM in total consumes ~30% of overall CH4 production. Our data corroborate a similar Q10 of AOM and methanogenesis. As the rate of AOM in paddy soils is lower than methanogenesis, however, it will not fully compensate for an increased methane production under climate warming.

Visualization and quantification of carbon "rusty sink" by rice root iron plaque: Mechanisms, functions, and global implications.

Paddies contain 78% higher organic carbon (C) stocks than adjacent upland soils, and iron (Fe) plaque formation on rice roots is one of the mechanisms that traps C. The process sequence, extent and global relevance of this C stabilization mechanism under oxic/anoxic conditions remains unclear. We quantified and localized the contribution of Fe plaque to organic matter stabilization in a microoxic area (rice rhizosphere) and evaluated roles of this C trap for global C sequestration in paddy soils. Visualization and localization of pH by imaging with planar optodes, enzyme activities by zymography, and root exudation by 14 C imaging, as well as upscale modeling enabled linkage of three groups of rhizosphere processes that are responsible for C stabilization from the micro- (root) to the macro- (ecosystem) levels. The 14 C activity in soil (reflecting stabilization of rhizodeposits) with Fe2+ addition was 1.4-1.5 times higher than that in the control and phosphate addition soils. Perfect co-localization of the hotspots of ß-glucosidase activity (by zymography) with root exudation (14 C) showed that labile C and high enzyme activities were localized within Fe plaques. Fe2+ addition to soil and its microbial oxidation to Fe3+ by radial oxygen release from rice roots increased Fe plaque (Fe3+ ) formation by 1.7-2.5 times. The C amounts trapped by Fe plaque increased by 1.1 times after Fe2+ addition. Therefore, Fe plaque formed from amorphous and complex Fe (oxyhydr)oxides on the root surface act as a "rusty sink" for organic matter. Considering the area of coverage of paddy soils globally, upscaling by model revealed the radial oxygen loss from roots and bacterial Fe oxidation may trap up to 130 Mg C in Fe plaques per rice season. This represents an important annual surplus of new and stable C to the existing C pool under long-term rice cropping.

Alternating Wet-Dry Cycles Rather than Sulfate Fertilization Control Pathways of Methanogenesis and Methane Turnover in Rice Straw-Amended Paddy Soil.

Alternate wet-drying (AWD) and sulfate fertilization have been considered as effective methods for lowering CH4 emissions from paddy soils. However, there is a clear knowledge gap between field studies that focus on the quantification of emissions and laboratory studies that investigate mechanisms. To elucidate mechanisms of CH4 production and oxidation under field conditions, rice was planted in straw-amended mesocosms with or without sulfate fertilization under continuously flooded conditions (FL) or two wet-dry cycles. CO2 and CH4 concentrations in soil air and their natural C isotope compositions were measured at stem elongation, booting, and flowering stages. CH4 concentration reached 51 mg C L-1 at the flowering stage under FL, while it decreased to 0.04 mg C L-1 under AWD. Relative 13C enrichment in CH4 and depletion in CO2 under AWD indicated CH4 oxidation. Ample organic substrate supply may have reduced competition between sulfate-reducing bacteria and methanogenic archaea, and therefore, it explains the absence of a decrease in CH4 concentrations in sulfate treatments. 13C enrichment in CO2 over time (6 and 7‰ with and without sulfate fertilizers, respectively) under FL indicates continuous contribution of hydrogenotrophic methanogenesis to CH4 production with ongoing rice growth. Overall, AWD could more efficiently reduce CH4 production than sulfate fertilization in rice straw-amended paddy soils.

Ecosystem carbon response of an Arctic peatland to simulated permafrost thaw.

Permafrost peatlands are biogeochemical hot spots in the Arctic as they store vast amounts of carbon. Permafrost thaw could release part of these long-term immobile carbon stocks as the greenhouse gases (GHGs) carbon dioxide (CO2 ) and methane (CH4 ) to the atmosphere, but how much, at which time-span and as which gaseous carbon species is still highly uncertain. Here we assess the effect of permafrost thaw on GHG dynamics under different moisture and vegetation scenarios in a permafrost peatland. A novel experimental approach using intact plant-soil systems (mesocosms) allowed us to simulate permafrost thaw under near-natural conditions. We monitored GHG flux dynamics via high-resolution flow-through gas measurements, combined with detailed monitoring of soil GHG concentration dynamics, yielding insights into GHG production and consumption potential of individual soil layers. Thawing the upper 10-15 cm of permafrost under dry conditions increased CO2 emissions to the atmosphere (without vegetation: 0.74 ± 0.49 vs. 0.84 ± 0.60 g CO2 -C m-2  day-1 ; with vegetation: 1.20 ± 0.50 vs. 1.32 ± 0.60 g CO2 -C m-2  day-1 , mean ± SD, pre- and post-thaw, respectively). Radiocarbon dating (14 C) of respired CO2 , supported by an independent curve-fitting approach, showed a clear contribution (9%-27%) of old carbon to this enhanced post-thaw CO2 flux. Elevated concentrations of CO2 , CH4 , and dissolved organic carbon at depth indicated not just pulse emissions during the thawing process, but sustained decomposition and GHG production from thawed permafrost. Oxidation of CH4 in the peat column, however, prevented CH4 release to the atmosphere. Importantly, we show here that, under dry conditions, peatlands strengthen the permafrost-carbon feedback by adding to the atmospheric CO2 burden post-thaw. However, as long as the water table remains low, our results reveal a strong CH4 sink capacity in these types of Arctic ecosystems pre- and post-thaw, with the potential to compensate part of the permafrost CO2 losses over longer timescales.

Microbial strategies for phosphorus acquisition in rice paddies under contrasting water regimes: Multiple source tracing by 32P and 33P.

Microbial acquisition and utilization of organic and mineral phosphorus (P) sources in paddy soils are strongly dependent on redox environment and remain the key to understand P turnover and allocation for cell compound synthesis. Using double 32/33P labeling, we traced the P from three sources in a P-limited paddy soil: ferric iron-bound phosphate (Fe-P), wheat straw P (Straw-P), and soil P (Soil-P) in microbial biomass P (MBP) and phospholipids (Phospholipid-P) of individual microbial groups depending on water regimes: (i) continuous flooding or (ii) alternate wetting and drying. 32/33P labeling combined with phospholipid fatty acid analysis allowed to trace P utilization by functional microbial groups. Microbial P nutrition was mainly covered by Soil-P, whereas microorganisms preferred to take up P from mineralized Straw-P than from Fe-P dissolution. The main Straw-P mobilizing agents were Actinobacteria under alternating wetting and drying and other Gram-positive bacteria under continuous flooding. Actinobacteria and arbuscular mycorrhiza increased P incorporation into cell membranes by 1.4-5.8 times under alternate wetting and drying compared to continuous flooding. The Fe-P contribution to MBP was 4-5 times larger in bulk than in rooted soil because (i) rice roots outcompeted microorganisms for P uptake from Fe-P and (ii) rhizodeposits stimulated microbial activity, e.g. phosphomonoesterase production and Straw-P mineralization. Higher phosphomonoesterase activities during slow soil drying compensated for the decreased reductive dissolution of Fe-P. Concluding, microbial P acquisition strategies depend on (i) Soil-P, especially organic P, availability, (ii) the activity of phosphomonoesterases produced by microorganisms and roots, and (iii) P sources - all of which depend on the redox conditions. Maximizing legacy P utilization in the soil as a function of the water regime is one potential way to reduce competition between roots and microbes for P in rice cultivation.

Keep oxygen in check: An improved in-situ zymography approach for mapping anoxic hydrolytic enzyme activities in a paddy soil.

Paddy soils regularly experience redox oscillations during the wetting and draining stages, yet the effects of short-term presence of oxygen (O2) on in-situ microbial hotspots and enzyme activities in anoxic ecosystems remain unclear. To fill this knowledge gap, we applied soil zymography to localize hotspots and activities of phosphomonoesterase (PME), ß-glucosidase (BG), and leucine aminopeptidase (LAP) in three compartments of rice-planted rhizoboxes (top bulk, rooted, and bottom bulk paddy soil) under oxic (+O2) and anoxic (O2) conditions. Short-term (35 min) aeration decreased PME activity by 13-49 %, BG by 4-52 %, and LAP by 12-61 % as compared with O2 in three soil compartments. The percentage of hotspot area was higher by 3-110 % for PME, by 10-60 % for BG, and by 12-158 % for LAP under +O2 vs. O2 conditions depending on a rice growth stage. Irrespective of the aeration conditions, the rhizosphere extent of rice plants for three enzymes was generally greater under higher moisture conditions and at earlier growth stage. Higher O2 sensitivity for the tested enzymes at bottom bulk soil versus other compartments suggested that short-term aeration during conventional zymography may lead to underestimation of nutrient mobilization in subsoil compared to top bulk soil. The intolerance of anaerobic microorganisms against the toxicity of O2 in the cells and the shift of microbial metabolic pathways may explain such a short-term suppression by O2. Our findings, therefore, show that anoxic conditions and soil moisture should be kept during zymography and probably other in-situ soil imaging methods when studying anoxic systems.

A novel belowground in-situ gas labeling approach: CH4 oxidation in deep peat using passive diffusion chambers and 13C excess.

In-vitro incubation of environmental samples is a common approach to estimate CH4 oxidation potential. Here we developed and verified an in-situ method utilizing passive diffusion chambers (PDC, silicone tubes) to deliver 13C-labeled CH4 into peat for the determination of the CH4 oxidation potential based on 13C excess of CO2. To target CH4 oxidation under semi-aerobic and anaerobic conditions, we installed 20 PDCs (30 ml volume) below the water table in profiles from 35-cm to 2-m depths of a peatland in north-eastern Sweden in July 2017 using a peat auger. 13C-labeled CH4 was injected into PDCs through tubing twice during 12 days (day 0 and 6) and samples were collected at days 1, 3, 6, 8 and 11. Background (non-labeled) δ13C of CO2 ranged from -7.3 (35 cm) to +5.7‰ (200 cm) with depth. These δ13C values rose to +110 and + 204‰ after the CH4 injection. The estimated CH4-derived C in CO2 was the lowest at the bottom of the profile (0.3 µmol L-1), whereas the maximum was at 100 cm (6.1 µmol L-1) at five days after the second labeling. This corresponded to 1.5-7.2% of the total CH4 pool to be oxidized, depending on depth. This novel approach with belowground in-situ 13C labeling of gases demonstrated the suitability of tracing the transformations of these gases in soil depth by PDCs and for the first time verified the in-situ occurrence of a deep-peat CH4 oxidation.

Microbial iron reduction compensates for phosphorus limitation in paddy soils.

Limitation of rice growth by low phosphorus (P) availability is a widespread problem in tropical and subtropical soils because of the high content of iron (Fe) (oxyhydr)oxides. Ferric iron-bound P (Fe(III)-P) can serve as a P source in paddies after Fe(III) reduction to Fe(II) and corresponding H2PO4- release. However, the relevance of reductive dissolution of Fe(III)-P for plant and microbial P uptake is still an open question. To quantify this, 32P-labeled ferrihydrite (30.8 mg P kg-1) was added to paddy soil mesocosms with rice to trace the P uptake by microorganisms and plants after Fe(III) reduction. Nearly 2% of 32P was recovered in rice plants, contributing 12% of the total P content in rice shoots and roots after 33 days. In contrast, 32P recovery in microbial biomass decreased from 0.5% to 0.08% of 32P between 10 and 33 days after rice transplantation. Microbial biomass carbon (MBC) and dissolved organic C content decreased from day 10 to 33 by 8-54% and 68-77%, respectively, suggesting that the microbial-mediated Fe(III) reduction was C-limited. The much faster decrease of MBC in rooted (by 54%) vs. bulk soil (8-36%) reflects very fast microbial turnover in the rice rhizosphere (high C and oxygen inputs) resulting in the mineralization of the microbial necromass. In conclusion, Fe(III)-P can serve as small but a relevant P source for rice production and could partly compensate plant P demand. Therefore, the P fertilization strategies should consider the P mobilization from Fe (oxyhydr)oxides in flooded paddy soils during rice growth. An increase in C availability for microorganisms in the rhizosphere intensifies P mobilization, which is especially critical at early stages of rice growth.

To shake or not to shake: Silicone tube approach for incubation studies on CH4 oxidation in submerged soils.

Incubation experiments are the most common approach to measure methane (CH4) oxidation potential in soils from various ecosystems and land-use practices. However, the commonly used headspace CH4 injection into microcosms and the shaking of the soil slurry during incubation fully removes CH4 (soil-born) and O2 (air-born) gradients common in situ, and may also induce various errors and disturbances. As an alternative, we propose CH4 input into microcosm soils via a silicone tube located within the slurry. We hypothesized that (i) poor CH4 diffusion in slurry will be compensated by direct CH4 delivery into the slurry via a silicone tube and, consequently, (ii) shaking of microcosms can be substituted with the soil silicone tube CH4 injection. During a 29-day submerged paddy soil incubation, the highest net CH4 oxidation rate was 1.6 µg C g-1 dry soil h-1, measured between the 3rd and 7th day after injecting 13CH4 into the slurry via a silicone tube without shaking. This rate was 1.5-2.5 times faster than the respective CH4 oxidation after headspace injection without shaking (1st hypothesis supported). As expected, shaking accelerated CH4 oxidation regardless of injection methods by 3.2-3.7 times (most intensively on days 3-7) compared to headspace injection without shaking. Nonetheless, the rates were similar between silicone tube injection without shaking and headspace injection with shaking. This supports the hypothesized potential of silicone tubes to substitute the common shaking method (2nd hypothesis). Furthermore, shaking increased the incorporation of 13C from CH4 into soil organic matter and microbial biomass by 1.8-2.7 times compared with CH4 injection into tubes and the static control without tubes. This reflects an overestimation of CH4 oxidation due to shaking. We conclude that direct soil CH4 injection via silicone tubes is advantageous in incubation experiments because gas concentration gradients are maintained and thereby more realistically reflect natural soil conditions.

Thermal stability of soil organic matter pools and their turnover times calculated by delta(13)C under elevated CO(2) and two levels of N fertilisation.

Soil from Free-Air Carbon dioxide Enrichment (FACE) plots (FAL, Braunschweig) under ambient air (375 ppm; delta(13)C-CO(2)-9.8 per thousand) and elevated CO(2) (550 ppm; for six years; delta(13)C-CO(2)-23 per thousand), either under 100% nitrogen (N) (180 kg ha(-1)) or 50% N (90 kg ha(-1)) fertilisation treatments, was analysed by thermogravimetry. Soil samples were heated up to the respective temperatures and the remaining soil was analysed for delta(13)C and delta(15)N by Isotope Ratio Mass Spectrometry (IRMS). Based on differential weight losses, four temperature intervals were distinguished. Weight losses in the temperature range 20-200 degrees C were connected mostly with water volatilisation. The maximum weight losses and carbon (C) content were measured in the soil organic matter (SOM) pool decomposed at 200-360 degrees C. The largest amount of N was detected in SOM pools decomposed at 200-360 degrees C and 360-500 degrees C. In all temperature ranges, the delta(13)C values of SOM pools were significantly more negative under elevated CO(2) versus ambient CO(2). The incorporation of new C into SOM pools was not inversely proportional to its thermal stability. 50% N fertilisation treatment gained higher C exchange under elevated CO(2) in the thermally labile SOM pool (200-360 degrees C), whereas 100% N treatment induced higher C turnover in the thermally stable SOM pools (360-500 degrees C, 500-1000 degrees C). Mean Residence Time of SOM under 100% N and 50% N fertilisation showed no dependence between SOM pools isolated by increasing temperature of heating and the renovation of organic C in those SOM pools. Thus, the separation of SOM based on its thermal stability was not sufficient to reveal pools with contrasting turnover rates of C.

CH4 and CO2 production below two contrasting peatland micro-relief forms: An inhibitor and δ13C study.

Two peatland micro-relief forms (microforms) - hummocks and hollows - differ by their hydrological characteristics (water table level, i.e. oxic-anoxic conditions) and vegetation communities. We studied the CH4 and CO2 production potential and the localization of methanogenic pathways in both hummocks and hollows at depths of 15, 50, 100, 150 and 200cm in a laboratory incubation experiment. For this purpose, we measured CH4 and CO2 production rates, peat elemental composition, as well as δ13C values of gases and solids; the specific inhibitor of methanogenesis BES (2-bromo-ethane sulfonate, 1mM) was aimed to preferentially block the acetoclastic pathway. The cumulative CH4 production of all depths was almost one fold higher in hollows than in hummocks, with no differences in CO2. With depth, CO2 and CH4 production decreased, and the relative contribution of the hydrogenotrophic pathway of methanogenesis increased. The highest methanogenic activity among all depths and both microforms was measured at 15cm of hollows (91%) at which the highest relative contribution of acetoclastic vs. hydrogenotrophic pathway (92 and 8%, respectively) was detected. For hummocks, the CH4 production was the highest at 50cm (82%), where relative contribution of acetoclastic methanogenesis comprised 89%. The addition of 1mM BES was not selective and inhibited both methanogenic pathways in the soil. Thus, BES was less efficient in partitioning the pathways compared with the δ13C signature. We conclude that the peat microforms - dry hummocks and wet hollows - play an important role for CH4 but not for CO2 production when the effects of living vegetation are excluded.

Stimulation of r- vs. K-selected microorganisms by elevated atmospheric CO(2) depends on soil aggregate size.

Increased root exudation under elevated atmospheric CO(2) and the contrasting environments in soil macro- and microaggregates could affect microbial growth strategies. We investigated the effect of elevated CO(2) on the contribution of fast- (r-strategists) and slow-growing (K-strategists) microorganisms in soil macro- and microaggregates. We fractionated the bulk soil from the ambient and elevated (for 5 years) CO(2) treatments of FACE-Hohenheim (Stuttgart) into large macro- (>2 mm), small macro- (0.25-2.00 mm), and microaggregates (<0.25 mm) using 'optimal moist' sieving. Microbial biomass (C(mic)), the maximum specific growth rate (mu), growing microbial biomass (GMB) and lag-period (t(lag)) were estimated by the kinetics of CO(2) emission from bulk soil and aggregates amended with glucose and nutrients. Although C(org) and C(mic) were unaffected by elevated CO(2), mu values were significantly higher under elevated than ambient CO(2) for bulk soil, small macroaggregates, and microaggregates. Substrate-induced respiratory response increased with decreasing aggregate size under both CO(2) treatments. Based on changes in mu, GMB and lag period, we conclude that elevated atmospheric CO(2) stimulated the r-selected microorganisms, especially in soil microaggregates. Such an increase in r-selected microorganisms indicates acceleration of available C mineralization in soil, which may counterbalance the additional C input by roots in soils in a future elevated atmospheric CO(2) environment.